In eukaryotic cells, transcription of gene is regulated, as the amount transcription, by interaction between sequences (cis-elements) on chromosomes such as promoters, enhancers, etc. and transcription factors binding thereto. A promoter is composed of the core (basic) sequence comprising TATA box and transcription start site, and the gene-specific expression regulatory region. Regulatory transcription factors bound to the expression regulatory region activate RNA polymerase II holoenzymes (complexes of RNA polymerase II and basal transcription factors) to promote gene transcription. The basal transcription factors bound to the core sequence are engaged in the transcription of many genes and are not specific to genes.
On the other hand, in regulatory transcription factors bound to the expression regulatory region, cells are activated when the cells proliferate or differentiate or respond to the external stimulations such as hormones, cytokines, etc., thus promoting the transcription of genes given. For this reason, regulatory transcription factors are also called gene-specific transcription factors. It is predicted that about 30,000 kinds of gene-specific transcription factors would be present in human and are expected to become drug targets, like receptors on cell surfaces (Mol. Med. Today, 358, 1998).
Human chondromodulin-I (hereinafter sometimes abbreviated as ChM-I) was purified and cloned from bovine fetal epiphyseal cartilage by Hiraki et al. (Biochem. Biophys. Res. Commun., 175:371, 1991), as a factor for promoting DNA synthesis of chondrocytes. Purified ChM-I weakly promoted DNA synthesis of chondrocytes and its activity was potentiated in the presence of bFGF. It was also shown that ChM-I promotes proteoglycan synthesis as well. It became clear from the cDNA sequence that ChM-I was synthesized as a precursor composed of 335, amino acids and mature type was composed of the C-terminal 121 amino acids. This mature ChM-I coincided with the 18-kDa glycoprotein, physiological functions of which are unknown and which P. J. Neame et al. (J. Biol. Chem., 265:9628-9633, 1990) purified from bovine septonasal cartilage and determined its partial amino acid sequence. It was also revealed by the Northern blot analysis that expression of bovine ChM-I was specific to cartilage. Subsequently, it is reported that ChM-I possesses a colony formation promoting activity of chondrocytes (Biochem. Biophys. Res. Commun., 241:395, 1997), a vascular endothelial cell growth inhibitory action (J. Biol. Chem., 272:32419, 1997, FEBS Letters, 415:321, 1997), a growth promoting/differentiation inhibitory action (FEBS Letters, 406:310, 1997), etc. In growth plate cartilages, ChM-I is expressed in the avascular proliferating cartilage zone, etc. It is thus considered that ChM-I would promote cartilage proliferation and matrix production, vascular invasion would be prevented, and the replacement of cartilage would be controlled by bone (J. Biol. Chem., 272:32419, 1997). As such, ChM-I is a factor for controlling the differentiation and proliferation of cartilage having an anti-angiogenic activity.
During endochondral ossification, chondrocytes in the growth plate cartilage continue to be differentiated in the order of resting cartilage, proliferating cartilage, hypertrophic cartilage and then calcified cartilage and, by vascular invasion, osteoblasts are supplied and then replaced by bone. Further in the course of healing bone fracture, chondrocytes appear after inflammatory reaction and bone fracture is restored via a similar pathway. Thus, the differentiation and proliferation of chondrocytes are clearly important for the process of healing osteopathy or chondropathy.
Suzuki et al. clarified the sequence of human ChM-I gene and described the application of human ChM-I protein to drugs for the treatment of bone fracture, various cartilage-associated diseases, tumors including cancer, etc., because human ChM-I protein possesses a cartilage proliferating action and vascular endothelial cell growth inhibition (Japanese Patent Laid-Open Application No. 7-138295). However, the controlled expression of ChM-I gene remains unclear not been made clear.
Regulatory transcription factors regulate the activity, expression and stability, and enable to develop novel pharmaceuticals useful for the prevention and treatment of various diseases associated with the activity, expression and stability, e.g., joint diseases such as arthritis deformans, chronic articular rheumatism, etc., osteopathy such as bone fracture, etc., and cancer, etc. It has thus been desired in the art to find a new regulatory transcription factor and develop a method of manufacturing the same in a large scale.